The mechanisms of the cooperative phenomena associated with the protein folding is not well understood. Understanding such phenomena is necessary to precisely design a functional protein. Thus, our studies are aimed at understanding these mechanisms. Previous studies have allowed us to assign 4 core domains in the cytochrome structure. A core domain is a structural region containing a hydrophobic core and surrounding shell which reversibly unfolds as a unit. Core domain 1 folds by itself and consists essentially of the amino and COOH-terminal helices and the heme (the top region). Core domains 2, 3 and 4 respectively, are assigned based on the core located at the left (the Fe -S bond) and right sides and at the bottom of the heme. Assembly of core domains 1, 2 and 3 forms the major part of the hydrophobic core surrounding the heme. To know whether the impact of mutation in this major part of the hydrophobic core in yeast iso-2 cytochrome c propagates itself, four core residues I20, M64, L85 and M98, one surface residues L9 and 4 exterior residues were mutated from residues found in iso-2 to those in horse cyt. c in previous years. The observations of current and previous years are: 1) Even a native mutation such as I20V can be deleterious in function depending on other residues in the core. Such a context dependency of deleterious mutational effect is reminiscent of the concept of the covarion of cyt. c evolution; 2) Almost all proton NMR resonances of the backbone chain of wild type iso-2 are assigned. The structures of M64L, M98L or M64L/M98L mutant iso-2 are close to that of wild type. Conformational mobility around the average structure increases in the order of wild type, M98L or M64L, M64L/M88L; 3) Small magnitude, non-additive mutational effects exist between L9I and M64L (long range) and between I20V and M98L (short range). Thus, we propose that the impact of mutation in the above 4 core and 1 surface residues propagates itself and that such a propagation is a manifestation of a process of re-establishment of the state of lowest free energy in a properly folded structural region as a single unit and that this structural region contains those formed by assembly of core domains 1,2 and 3. Surprisingly, transformation of a majority of residues in the major part of the core from iso-2 to horse cyt. c resulted in no increase in stability despite the much greater stability of wt horse cyt. c over that of wt iso-2. Previous studies suggest that core domains 1, 2 and 3 must be assembled to generate such a stability difference. Thus, to obtain a clue to the origin of this phenomenon, 2 types of chimeric cyts. c have been prepared: one contains iso-2 core and yeast iso-1 cyt. c shell plus 4 extra NH2 -terminal residues which exist only in iso-2 and the other, iso-1 core and iso-2 shell. Thermodynamic analysis of these chimera, C102A iso-1 and wild type iso-2 suggest that there could be unique core-shell interactions which modulates stability of cyt. c.